Page 195 - 2020_01-Haematologica-web

Basal NF-κB inhibition impacts BCR responsiveness in CLL
A
BC
DE
FG
Figure 1. Ca2+ signaling in chronic lymphocytic leukemia cells. (A) Flow cytometric analysis of Ca2+ flux (ratio Indo-1/Indo-1) after the addition of 4-hydroxytamoxifen (4-OHT) to triple knockout (TKO) cells expressing the B cell receptor (BCR) from two representative samples of mutated chronic lymphocytic leukemia (M-CLL) (left) and two unmutated samples (U-CLL) (right). (B) From nine CLL cases (6 U-CLL, black dots and 3 M-CLL, open dots) in whom the basal Ca2+ level (x-axis) had been assessed earlier, the BCR was cloned into TKO cells to determine the autonomous Ca2+ signal (y-axis). Linear regression was performed and the R2 and P-value are shown. (C) Basal Ca2+ level [median fluorescence intensity (MFI) ratio FI3/FR] was determined in a new cohort of 52 CLL samples (freshly isolated) consisting of 30 U-CLL and 22 M-CLL cases. (D) Responsive capacity upon BCR stimulation. Flow cytometric analysis of a representative CLL sample showing no Ca2+ influx (ratio FL3/FR) upon α-IgM stimulation (left) and a representative CLL sample with an increase in Ca2+ influx (ratio FL3/FR) upon α-IgM stimulation (right). Based on this analysis the responsive capacity upon α-IgM stimulation (E) and α-IgD stimulation (F) was determined in the 30 U-CLL and 22 M-CLL cases. Individual plots and medi- ans (gray bars) are shown. The Mann-Whitney U-test was performed for statistical analysis between the groups of patients (*P<0.05). (G) Linear regression analysis between the relative response after α-IgM and α-IgD stimulation: R2 and P values are shown.
haematologica | 2020; 105(1)
185